ABSTRACT
Central neural networks operate continuously throughout life to control respiration, yet mechanisms regulating ventilatory frequency are poorly understood. Inspiration is generated by the pre-Bötzinger complex of the ventrolateral medulla, where it is thought that excitation increases inspiratory frequency and inhibition causes apnea. To test this model, we used an in vitro optogenetic approach to stimulate select populations of hindbrain neurons and characterize how they modulate frequency. Unexpectedly, we found that inhibition was required for increases in frequency caused by stimulation of Phox2b-lineage, putative CO2-chemosensitive neurons. As a mechanistic explanation for inhibition-dependent increases in frequency, we found that phasic stimulation of inhibitory neurons can increase inspiratory frequency via postinhibitory rebound. We present evidence that Phox2b-mediated increases in frequency are caused by rebound excitation following an inhibitory synaptic volley relayed by expiration. Thus, although it is widely thought that inhibition between inspiration and expiration simply prevents activity in the antagonistic phase, we instead propose a model whereby inhibitory coupling via postinhibitory rebound excitation actually generates fast modes of inspiration.
Subject(s)
Carbon Dioxide/pharmacology , Exhalation/drug effects , Inhalation/drug effects , Neurons/drug effects , Respiratory Center/drug effects , Respiratory Rate/drug effects , Animals , Carbon Dioxide/metabolism , Exhalation/physiology , Female , Hypoglossal Nerve/drug effects , Inhalation/physiology , Male , Medulla Oblongata/cytology , Medulla Oblongata/drug effects , Medulla Oblongata/physiology , Mice , Neurons/cytology , Neurons/physiology , Optogenetics/methods , Phrenic Nerve/drug effects , Picrotoxin/pharmacology , Prazosin/pharmacology , Propranolol/pharmacology , Respiratory Center/cytology , Respiratory Center/physiology , Respiratory Rate/physiology , Spinal Nerve Roots/drug effects , Strychnine/pharmacology , Substance P/pharmacologyABSTRACT
Spinal cord injury (SCI) above cervical level 4 disrupts descending axons from the medulla that innervate phrenic motor neurons, causing permanent paralysis of the diaphragm. Using an ex vivo preparation in neonatal mice, we have identified an excitatory spinal network that can direct phrenic motor bursting in the absence of medullary input. After complete cervical SCI, blockade of fast inhibitory synaptic transmission caused spontaneous, bilaterally coordinated phrenic bursting. Here, spinal cord glutamatergic neurons were both sufficient and necessary for the induction of phrenic bursts. Direct stimulation of phrenic motor neurons was insufficient to evoke burst activity. Transection and pharmacological manipulations showed that this spinal network acts independently of medullary circuits that normally generate inspiration, suggesting a distinct non-respiratory function. We further show that this "latent" network can be harnessed to restore diaphragm function after high cervical SCI in adult mice and rats.
